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Double Hit Boosts Neuron Production

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Review of “Combining Suppression of Stemness with Lineage-Specific Induction Leads to Conversion of Pluripotent Cells into Functional Neurons” from Chemistry & Biology by Stuart P. Atkinson.

The differentiation of pluripotent stem cells (PSCs) into therapeutically relevant somatic cell types may provide the means to treat various debilitating diseases and disorders. However, current protocols suffer from low efficiency so creating a considerable barrier to their voyage from the bench to the bedside.

To get round this barrier, researchers from the laboratories of Kyeong Kyu Kim and Injae Shin hoped to strike twice at PSCs to boost the production of functional neurons. To do this, they aimed to inhibit the pluripotent state and promote neural differentiation through the application of synthetic neurogenic inducers in the hope that this may increase differentiation efficiency. This new study, published in Chemistry & Biology, represents an interesting new strategy that may be applicable to a wide range of differentiation techniques [1].

To inhibit pluripotency, the authors utilized an engineered bacterial molecular chaperone protein (Skp) known to inhibit Sox2 activity and enhance neuronal/cardiac differentiation [2] in the pluripotent P19 embryonic carcinoma cell line. The addition of a TAT cell-penetrating peptide and a nuclear localization signal (NLS) allowed cell transduced Skp to downregulate stemness-related genes and also upregulate lineage-specific differentiation genes in P19 cells cultured as embryoid bodies.

To boost neurogenic differentiation, the authors then cultured the differentiating P19 cells in a monolayer in the presence of synthetic neurogenic inducers (neurodazine [Nz] or neurodazole [Nzl]) [3]. This specifically enhanced neuronal differentiation, as compared to P19 cells not treated with Skp, as measured by the abundance of the pan-neuronal marker Tuj1. Transcriptional analysis of the resultant cells demonstrated an upregulation of genes related to the construction of several sodium channels suggesting that they may be electrophysiologically active and, therefore, functional. Encouragingly, functionality was then confirmed through whole-cell patch-clamp analyses which demonstrated fast-inactivating inward and outward currents in around 70% of cells.

This double hit to inhibit pluripotency and promote neurogenesis seems to be an effective strategy and this will success will hopefully be extended from embryonic carcinoma cells to the more clinically relevant embryonic stem cells and induced pluripotent stem cells. Furthermore, Skp may also enhance cardiac cell differentiation, and so the combination of Skp with known cardiac enhancers may also be a fruitful line of investigation.

References

  1. Halder D, Chang GE, De D, et al. Combining Suppression of Stemness with Lineage-Specific Induction Leads to Conversion of Pluripotent Cells into Functional Neurons. Chem Biol 2015;22:1512-1520.
  2. De D, Jeong MH, Leem YE, et al. Inhibition of master transcription factors in pluripotent cells induces early stage differentiation. Proc Natl Acad Sci U S A 2014;111:1778-1783.
  3. Kim GH, Halder D, Park J, et al. Imidazole-based small molecules that promote neurogenesis in pluripotent cells. Angew Chem Int Ed Engl 2014;53:9271-9274.